The production of antibodies by antibody-secreting cells (ASCs) is critically important for immune responses to many different pathogens. However, in autoimmune diseases where B cell tolerance is broken, many ASCs secrete autoantibodies and contribute in a major way to organ pathology. The differentiation of B cells into ASCs is controlled by a cohort of key transcription factors including Ets-1, which serves as a negative regulator of this process. In the absence of Ets-1, B cells undergo enhanced differentiation into ASCs many of which secrete autoantibodies as demonstrated by high titers of IgM and IgG autoantibodies in the serum. Recently Toll- like receptor (TLR) signaling, particularly via TLR7 and TLR9, has been implicated in the activation of autoreactive B cells to differentiate into ASCs. Interestingly, Ets-1 deficient B cells exhibit enhanced responses to TLR7 and TLR9 ligands in vitro and Ets-1 knockout mice lacking the TLR adaptor protein Myd88 have reduced autoantibody production. Toll-like receptor signaling is a potent inducer of the expression of Blimp-1, a key transcription factor that drives ASC differentiation. Ets-1 physically interacts with Blimp-1 to inhibit the ability of Blimp-1 to bind target DNA sequences. In contrast, a closely related transcription factor Ets-2 is unable to block Blimp-1 binding or to inhibit ASC differentiation. Ets-1 also is thought to regulate the expression of critical target genes controlling ASC formation, but the identity of most of these targets remains unclear. In this application we propose a variety of assays to further define the role of Ets-1 in regulating target gene expression, Blimp-1 activity and ASC differentiation. These studies will be aided by comparing biological activities of Ets-1, which can block ASC differentiation, with the closely related Ets-2 protein, which lacks this activity. The specific aims of the proposal are (1) to determine which cell types require Ets-1 activity to limit ASC formation, autoantibody secretion and autoimmune disease, (2) to examine the in vivo role of TLR7 and 9 ligands in activating Ets-1 deficient B cells, (3) to identify Ets-1 dependent gene expression pathways regulating ASC formation and (4) to identify structural features of Ets-1 that impart a unique ability to regulate ASC development. Together our studies will provide novel insights into the transcriptional regulation of B cell differentiation particularly in situations of autoantibody secretion. PUBLIC HEALTH RELEVANCE: As part of the immune response to pathogens, B cells differentiate into antibody-secreting cells (ASCs) that produce high levels of antibodies to fight infection. In autoimmune disease, there are large numbers of ASCs that secrete antibodies that react with self-tissues. Information gleaned from the studies proposed will help define the molecular details of how B cells differentiate into ASCs and how that process is defective in autoimmune disease, thereby potentially identifying new strategies to stimulate or inhibit this process clinically.